The alarmin interleukin-1α triggers secondary degeneration through reactive astrocytes and endothelium after spinal cord injury

Spinal cord injury (SCI) triggers neuroinflammation, and subsequently secondary degeneration and oligodendrocyte (OL) death. We report that the alarmin interleukin (IL)−1α is produced by damaged microglia after SCI. Intra-cisterna magna injection of IL-1α in mice rapidly induces neutrophil infiltration and OL death throughout the spinal cord, mimicking the injury cascade seen in SCI sites. These effects are abolished through co-treatment with the IL-1R1 antagonist anakinra, as well as in IL-1R1-knockout mice which demonstrate enhanced locomotor recovery after SCI. Conditional restoration of IL-1R1 expression in astrocytes or endothelial cells (ECs), but not in OLs or microglia, restores IL-1α-induced effects, while astrocyte- or EC-specific Il1r1 deletion reduces OL loss. Conditioned medium derived from IL-1α-stimulated astrocytes results in toxicity for OLs; further, IL-1α-stimulated astrocytes generate reactive oxygen species (ROS), and blocking ROS production in IL-1α-treated or SCI mice prevented OL loss. Thus, after SCI, microglia release IL-1α, inducing astrocyte- and EC-mediated OL degeneration.


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We require information from authors about some types of materials, experimental systems and methods used in many studies. Here, indicate whether each material, system or method listed is relevant to your study. If you are not sure if a list item applies to your research, read the appropriate section before selecting a response. Statistical evaluations were performed using the Student's t-test or one-or two-way ANOVA or repeated-measures ANOVA, as indicated in the figure legends. Post-ANOVA comparisons were made using the Bonferroni correction. For behavioral analyses, the number of mice per group was >8, which represents a >98% probability of detecting a significant change if alpha is set at 0.05 and standard deviations (SDs) are 20% of average. Histological and immunohistochemical comparisons were performed using unpaired Student's t-test (2 groups) or a one-way (>2 groups) ANOVA, followed by post hoc correction. At least 4 mice per condition were used. This represents a >98% probability of detecting a significant change if alpha is set at 0.05 and SDs are 8% of average. Statistical powers were calculated using the maximum SD observed in our previous studies. For the in vitro experiments, the values of 6 wells were averaged for each tested condition.
No data were excluded from the analyses.
Data were pooled from two independent experiments for the following figures: Fig Negative control groups (e.g. genotype, vehicle treatment) were included in all experiments. Male and female mice were randomly assigned (in equal numbers) to control and treatment groups.
All quantifications were done blind with respect to the identity of the animals. Behavioral testing was carried out by a blinded investigator.

Flow Cytometry
Plots Confirm that: The axis labels state the marker and fluorochrome used (e.g. CD4-FITC).
The axis scales are clearly visible. Include numbers along axes only for bottom left plot of group (a 'group' is an analysis of identical markers).
All plots are contour plots with outliers or pseudocolor plots.
A numerical value for number of cells or percentage (with statistics) is provided.
The antibodies used in this study were chosen based on their extensive use in the literature (supported by the references available on the supplier's website) and a significant amount of experiments in our hands (see the references provided in the Methods section).
Oligodendrocyte precursor cells (OPCs) were isolated from the neonatal (P7-P9) mouse brain. Endothelial cells were isolated from the brain capillaries of mice aged 6-8 weeks. Mouse primary astrocytes were isolated from the cortex of P0-P2 C57BL/6 mice.
Morphology and immunophenotypic characterization were performed to assess the purity of the cultures.
The cell lines were not tested for mycoplasma contamination.

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Male and/or female C57BL/6 mice were purchased from Charles River Laboratories or The Jackson Laboratory (JAX) at 8-10 weeks of age. Cx3cr1CreER mice were obtained from the European Mouse Mutant Archive, with prior authorization from Dr. Steffen Jung (Rehovot, Israel). Breeders for Rosa26-tdTomato (R26-TdT, also known as Ai9, stock #007905), PdgfraCreER (stock #018280), GfapCre (stock #024098), and Il1r1fl/fl (stock #028398) mice were all purchased from JAX. LysM-eGFP knock-in mice were obtained from Dr. Gregory Dekaban (Robarts Institute, London, ON, Canada), with prior consent of Dr. Thomas Graf (Center for Genomic Regulation, Barcelona, Spain). Cdh5CreER mice (line #13073), in which the tamoxifen-inducible Cre recombinase is active in all ECs, were purchased from the Cancer Research Technology Repository at Taconic with prior consent of the creator of the mouse line, Dr. Ralf Adams (London Research Institute, UK). Il1r1r/r mice were obtained from Dr. Ning Quan. Ly6gCre-TdT mice were provided by Dr. Matthias Gunzer (University of Duisburg-Essen, Essen, Germany). All transgenic mouse lines are described in more detail in the manuscript and references provided in it. Mice were housed in individually ventilated cages on racks connected to a central HEPA filtered air supply (30-70 air changes per hour). All animals were kept in a standard 12-h light-dark cycle and had free access to food and water at all times. Room temperature was maintained at 23 ± 2°C with a relative humidity of 50 ± 5%.
No wild animals were used in the study.
No field collected samples were used in the study.
All animal procedures were approved by the Comité de protection des animaux de l'Université Laval (protocols #CHU-20-675 and #CHU-21-860) and conducted in compliance with relevant ethical regulations and guidelines of the Canadian Council on Animal Care. AUTOMATED BLOOD CELL COUNT, FLOW CYTOMETRY AND CELL SORTING. Blood was collected via cardiac puncture using a